A promising concept for extraction and solubilization of membrane proteins, particularly hydrophobic proteins, is the use of a detergent. Detergents are amphipathic molecules, which contain polar and non-polar chemical groups. Consequently, they exhibit unique properties in water. They are soluble in water and can solubilize hydrophobic proteins by interacting with hydrophobic domains (for example transmembrane regions). Numerous detergents are known to the public domain but in principle they can be divided into non-ionic, ionic and zwitterionic detergents. The solubilizing potency of detergents varies depending on the hydrophilic/lipophilic balance (HLB) of the amphiphilic groups in the molecule. Detergents with high solubilization power (like Sodium Dodecyl Sulfate—SDS) have also denaturing effects on the structure of the proteins to be solubilized. The selection of the appropriate detergent for the intended use is therefore dependent on the properties of the target protein and the technical conditions of the process (see, e.g., L. M. Hjelmeland and A. Chrambach, “Solubilization of Functional Membrane Proteins”, Meth. Enzymol., 1984, Vol. 104, Part C, pages 305-328).
One of the major disadvantages of using detergents for solubilizing biomolecules such as proteins is the contamination of the desired biomolecule with the detergent itself. Complete removal of the detergent is generally time-consuming and tedious, or even impossible in some cases. Further, by requiring high number of post-extraction purification steps the resulting overall yield of the desired product can decrease to an uneconomical degree.
Further, the most common method known in the art for cell debris separation is centrifugation. However, centrifugation, especially in view of an industrial large production scale process, shows a variety of drawbacks, such as high cost of industrial scale centrifuges and low efficacy in case of fine particles.
Thus, a strong need exists for a method useable in obtaining a highly purified hydrophobic protein from cells which overcome the above-mentioned disadvantages.
Therefore, it is an object of the present invention to provide a new method for obtaining highly purified proteins from cells, such as membrane proteins, particularly hydrophobic proteins.